Extremity trauma causing peripheral nervous system (PNS) injury accounts for the majority of combat wounds. Despite high regenerative capacity of the PNS, patients develop severe neuropathic pain, not amenable to analgesic therapies. The congressional Opioids and STOP Initiative Act of 2017 calls to ?expand, intensify, and coordinate fundamental, translational, and clinical research? on pain and develop new non-addictive pain treatments. In addition to physical and emotional disability, chronic pain costs the U.S. over $ 600 billion every year. Over 65% of American Veterans report pain, with severe pain 40% greater in Veterans than non- Veterans. Our unbiased transcriptomics study identified Tissue Inhibitor of Metalloproteinases-1 (TIMP-1) among the top-10 induced (out of thousands regulated) genes in painful PNS injury. The main function of TIMP-1 is inhibition of the matrix metalloproteinase (MMP) family of extracellular proteases. In the framework of this VA Merit program, we have pioneered the study of the MMP/TIMP axis in PNS injury and pain regulation. This renewal application centers on our second unbiased finding: TIMP-1 is an X-chromosome-linked gene, exhibiting polymorphic, aberrant and sex-dependent transcript isoforms in painful PNS injury. Our data strongly suggest that TIMP-1 is an analgesic, pro-survival and regenerative factor induced in the PNS in response to injury. However, expression of polymorphic, aberrant TIMP-1 transcript variants, potentially resulting in dysfunctional gene or protein products, predisposes to chronic pain development. With our innovative tools and concepts, groundbreaking data and strong track record in MMP/TIMP and PNS research, we aim to study cellular and subcellular, including nuclear patterns of TIMP-1 distribution, interactors and functions in PNS injury (Aim 1). We will then identify sex-specific, aberrant, transcript variants of TIMP-1 gene arising due to sexually dimorphic (X chromosome inactivation in females) and monomorphic (universal in both sexes) epigenetic abnormalities (Aim 2). As polymorphic TIMP-1 variants occur in humans, we expect our findings will swiftly translate into medical epigenetic diagnostics of pain states in a clinical setting. Finally we aim to develop targeted TIMP-1 gene therapy in PNS injury and pain (Aim 3). This program employs multidisciplinary state-of-the-art (e.g. RNA-seq, SMRT-BS, BioID, ChiP) technologies and fundamental neuropathological and behavioral approaches to study PNS injury and pain. We anticipate our program will make a major impact on pain diagnostics and development of non-addictive analgesics.